Articular cartilage degeneration in osteoarthritis is a major cause of disability affecting more than 43 million lives in the US. Damaged cartilage does not heal. Cartilage tissue engineering based on mesenchymal stem cell (MSC) technology offers the promise of manufacturable autologous tissue replacements of arbitrary size and shape, using cells from a bone marrow biopsy. This Bioengineering Research Grant proposal is driven by the need for cartilage tissue engineering to make the transition to a reproducible clinical therapy. To accomplish this, the processes involved must be thoroughly understood, and standardized protocols must be established. The global hypothesis underlying this proposal is that by exposing the cell scaffold constructs in a controlled, in vitro environment to conditions that favor chondrogenic differentiation, these constructs will develop the specific properties required for survival after implantation in the joint. Two major problems must be solved before tissue engineering can become a routine treatment for cartilage defects: transport of nutrients and waste products to and from the cells within the construct, and mechanical conditioning of the constructs to allow function in situ. We propose five Specific Aims, which address specific issues related to mass transport and mechanical conditioning, and their impact on construct functionality in vivo. These are: Specific Aim 1: To assess the mass transfer problem in cartilage tissue engineering. This will lead to better understanding of mass transfer within the constructs and will serve to evaluate the counter-measures proposed to improve mass transfer. Specific Aim 2: To develop a cartilage bioreactor monitoring and process control system. We will implement this technology to identify and characterize process control parameters in cartilage tissue engineering systems. Specific Aim 3: To develop countermeasures to specific mass transport limitations. In this Specific Aim, we will examine strategies to improve mass transfer in the constructs. Specific Aim 4: To implement and assess mechanical stimulation of the composites, using a system in which controlled, physiologically relevant loads can be applied, and identifying loading parameters that improve the mechanical properties of the construct. Specific Aim 5: To evaluate the composite constructs in vivo, as survival and integration of the manufactured construct in the joint defines the success of the tissue engineering process. The practical value of the proposed research is in the new insights it will provide into the technical issues surrounding cartilage tissue engineering and cartilage repair, and into the complex biology of the joint. If successful, these studies will provide novel principles and guidelines for the successful management of articular cartilage injuries.